A fundamental aspect of cellular signaling is the release of Ca from sequestered intracellular stores in the sarcoplasmic reticulum (SR) or endoplasmic reticulum (ER). For efficient operation of excitation-contraction coupling in muscle cells, or excitation-secretion coupling in non-muscle cells, counter ion movement across the SR or ER membrane must accompany the rapid efflux of Ca to neutralize the transient negative potential produced during Ca release. Although channels selective for monovalent cations have been reported in SR or ER membranes, no gene has been identified that encodes a SR or ER localized cation selective channel. We have recently discovered a novel gene encoding a trimeric intracellular cation-selective channel (TRIC) that contains an intracellular membrane-retention sequence at the carboxyl-terminus and a conserved leucine-zipper motif at the amino-terminus. Purified TRIC protein can form a cation-selective channel in lipid bilayer membrane, providing the possibility that TRIC may provide certain aspects of the counter-ion movement mechanism during the acute phase of Ca release. Biochemical studies identify a functional interaction between TRIC and SERCA, which may influence Ca homeostasis inside the SR. The focus of this project is to define the cellular and molecular functions of TRIC as a counter-ion channel in regulating the permeability of SR membrane to monovalent cations and/or as a modulator of Ca transport across the SR membrane. We will first establish the single channel properties of TRIC using the lipid bilayer reconstitution system. Through biochemical and molecular assays, we will identify the interacting partners for TRIC that may participate in regulating the channel activity of TRIC, or in modulating the functional interaction between TRIC and SERCA in skeletal muscle. Through comparative studies between wild type control and mutant muscle fibers lacking TRIC, we will define the physiological role of TRIC in mediating both the acute phase of Ca release from the SR and active Ca uptake into the SR. As TRIC represents a novel class of cation channels targeted to an intracellular organelle, knowledge of the role of this protein in muscle physiology will provide us valuable insights into the regulation of Ca signaling in muscle-related diseases, and in other diseases where dysfunctional Ca regulation results in pathology, such as neurodegenerative disorders. [unreadable] [unreadable]